MALACHITE GREEN: CHARACTERISTICS, PREPARATION, APPLICATIONS, TOXICITY - BIOLOGY - 2024

The malachite green is a dye of organic nature having a beautiful very similar to malachite green mineral, hence its name arises. However, malachite green dye and the mineral from which it takes its name are not related to each other.

It is used as a stain in various procedures, including spore staining (Shaeffer – Fulton or Wirtz-Conklin) and in mounting stool samples using the Kato concentration technique.

Malachite green crystals. Source: W. Oelen

Malachite green was formerly used as an antiparasitic in the treatment of captive-bred freshwater fish (aquarium and fish tanks). These fish can often be affected by protozoa such as Ichthyophthirius multifiliis or Dactylogyrus vastator.

Due to its bactericidal and fungicidal effect, malachite green dye is used as an inhibitor of the commensal microbiota in selective culture media for mycobacteria (Löwenstein-Jensen).

However, it has been seen that malachite green does not biodegrade easily, remaining in the environment in a residual form for a long time in aquatic foods. This affects the life of certain types of fish, crustaceans, and aquatic microorganisms.

On the other hand, malachite green has a reduced form that appears under anaerobic conditions called leukomalachite green, which is even more toxic. For this reason, today the use of malachite green is in disuse and in some countries it has been prohibited.

characteristics

Malachite green is a dark green crystalline-looking powder. The zinc-free presentation is preferred because it is less harmful, especially when used to treat fish.

It is also known as malachite oxalate green, aniline green, basic green 4, diamond green B, and Victoria green B. Its chemical formula is C ₅₂ H ₅₄ N ₄ O _12. Its scientific name is Chloride of 4- {(phenyl) methylidene} -N, N-dimethylcyclohexa-2,5-diene-1-iminium.

It can be found in the form of the oxalate salt or in the form of chloride. It has a solubility in water of 20 g / L ^-1, and it is also soluble in methanol. Its molecular mass is 369.91 g / mol ^-1. Its ionization constant is 6.90 pKa.

When in solution it is characterized by having two mixed forms (the cationic one at acidic pH) and its basic form (carbinol). Therefore, depending on the pH, one or the other will be favored.

Preparation

Malachite green for colorations

- Weigh 5 grams of malachite green.

- Dissolve in 100 ml of water.

Malachite green for the Kato-Katz

For this test, 3% malachite green is used

Applications

Shaeffer – Fulton spore staining

This staining technique, also known as Wirtz-Conklin, uses malachite green to stain the spores inside and outside the cell and safranin to counteract the vegetative cell. Therefore, the spore is stained green and the vegetative cell red.

Helminth concentration test (Kato-Katz)

This method is used for the diagnosis of helminth parasitosis. The technique is based on the use of glycerin as a lightener and malachite green as a contrast (it reduces the brightness of the sample).

However, the malachite green in this technique is not essential. If this dye is not available, it can be done without it.

Preparation of culture media

Due to the bactericidal and fungicidal effect that malachite green has, it is used in the preparation of the Löwenstein-Jensen culture medium, a special medium for the isolation of mycobacteria.

In the food and textile industry

For some time it was used as a colorant to dye paper and fabrics, but its use increased the toxic power of industrial waste that is dumped into the sea. This has produced the contamination of marine foods, such as fish, prawns, among others.

On the other hand, the use of malachite green in fish farms for the treatment of parasitosis in fish can lead to marine products for human consumption with the presence of the dye. This is serious due to its carcinogenic effect. As a result, it is no longer used in these areas.

Bacterial plaque staining

Various chemicals have been used throughout history as a method for visualizing bacterial plaque at the dental level. Among them is malachite green, along with others such as erythrosine.

Treatment of fish and crustaceans in ponds, fish tanks and aquariums

Malachite green is useful for the treatment of fish infested with the ciliate protozoan Ichthyophthirius multifiliis. This parasite causes white spot disease. This parasitosis is a real problem for those who breed and keep fish in captivity, due to its easy spread.

Malachite green is used in a wide variety of ways to treat affected fish. For example, it has been used by giving the fish a short bath, transferring them to a container with water and malachite green.

Another way is by pouring a specific amount of this substance into the water or by placing malachite green directly on the animal's skin.

Disease prevention in fish

Not only can fish diseases be treated with this compound, but it is also possible to prevent it. New fish entering the tank can be disinfected, passing the specimens through special baths that contain malachite green as a disinfectant, among other substances.

Fish eggs can also be disinfected with malachite green solution before being transferred to other ponds.

Considerations for treating fish and crustaceans with malachite green

Although malachite green eliminates some diseases, it is also true that it is harmful to bacteria and saprophytic fungi and to some types of fish, affecting respiratory enzymes. It also remains in a residual form in aquatic environments.

Therefore, its use is not currently recommended. The use of malachite green is banned in many countries, including the United States, the European Union, and some Southeast Asian countries such as Thailand.

Toxicity

Health effects

The NFPA (National Fire Protection Association) classifies malachite green as follows:

- Health risk in grade 2 (moderate risk).

- Risk of flammability in grade 1 (slight risk).

- Reactivity risk in grade 0 (no risk).

The health risks are irritation of the skin and mucous membranes from direct contact. It can become serious on the ocular mucosa.

It also causes irritability of the respiratory and digestive tracts by inhalation and accidental ingestion, respectively.

The consumption of contaminated food has a high carcinogenic power.

Impact on the environment

Malachite green has been shown to generate long-lasting residual harmful effects in aquatic environments, affecting beneficial saprophytic microbiota and some types of fish. For this reason the remnants of this colorant should not be poured down the drain.

In this sense, Iannacone and Alvariño studied the aquatic ecotoxicity of two colorants (methylene blue and malachite green) on Daphnia magna (crustacean). They concluded that both dyes are toxic, although methylene blue is to a greater extent. Both affect the aquatic environment at concentrations less than 1%.

On the other hand, various investigations have revealed that the toxicity of malachite green is influenced by temperature and pH. In this sense, the toxicity is higher with increasing temperature and decreasing pH.

Due to the above, the FDA (Food and Drug Administration) banned the use of malachite green in aquaculture in 1991, due to its carcinogenic properties

Incompatibility

Malachite green is recommended not to be used with zinc and also not to contact iron. It should be stored away from oxidizing substances.

References

1. Rabbit R. Kato method (Parasitological Diagnosis and its use as a Field Work Instrument). Medical Journal of Costa Rica. 1976; 43 (457) 169-174. Available at: binasss.sa.cr/revistas/rmcc.
2. Malachite green. Wikipedia, The Free Encyclopedia. 25 Dec 2018, 22:34 UTC. 27 May 2019, 20:30 en.wikipedia.org
3. National Institute of Rehabilitation. Malachite green. Biosafety Committee Safety Data Sheets. Available at: inr.gob.mx
4. Pontifical Javeriana University. 2015. Malachite green. Available: javeriana.edu.co/
5. Meyer Chemistry Reagents. 2009. Malachite green. Safety data Sheet. Available at: reactivosmeyer.com
6. Iannacone J, Alvariño L. Aquatic ecotoxicity of two dyes and of three important antiparasitics in aquaculture in Daphnia magna. Ecol. Apl. 2007; 6 (1-2): 101-110. Available at: Scielo.org.
7. López N. Determination of food coloring in food products from the sea. 2011. Degree work presented to obtain the Official University Master's Degree: Pesticide and pollutant residues. Food and environmental control. University of Almeira. Available at: repository.ual.es/bitstream.